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1.
Genes Dev ; 33(15-16): 936-959, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31123059

RESUMO

Changes in chromatin structure mediated by ATP-dependent nucleosome remodelers and histone modifying enzymes are integral to the process of gene regulation. Here, we review the roles of the SWI/SNF (switch/sucrose nonfermenting) and NuRD (nucleosome remodeling and deacetylase) and the Polycomb system in chromatin regulation and cancer. First, we discuss the basic molecular mechanism of nucleosome remodeling, and how this controls gene transcription. Next, we provide an overview of the functional organization and biochemical activities of SWI/SNF, NuRD, and Polycomb complexes. We describe how, in metazoans, the balance of these activities is central to the proper regulation of gene expression and cellular identity during development. Whereas SWI/SNF counteracts Polycomb, NuRD facilitates Polycomb repression on chromatin. Finally, we discuss how disruptions of this regulatory equilibrium contribute to oncogenesis, and how new insights into the biological functions of remodelers and Polycombs are opening avenues for therapeutic interventions on a broad range of cancer types.


Assuntos
Montagem e Desmontagem da Cromatina/genética , Proteínas Cromossômicas não Histona/metabolismo , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/metabolismo , Neoplasias/fisiopatologia , Proteínas do Grupo Polycomb/metabolismo , Fatores de Transcrição/metabolismo , Animais , Regulação da Expressão Gênica no Desenvolvimento , Humanos
2.
RNA ; 25(8): 1004-1019, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31097619

RESUMO

The marsupial inactive X chromosome expresses a long noncoding RNA (lncRNA) called Rsx that has been proposed to be the functional analog of eutherian Xist Despite the possibility that Xist and Rsx encode related functions, the two lncRNAs harbor no linear sequence similarity. However, both lncRNAs harbor domains of tandemly repeated sequence. In Xist, these repeat domains are known to be critical for function. Using k-mer based comparison, we show that the repeat domains of Xist and Rsx unexpectedly partition into two major clusters that each harbor substantial levels of nonlinear sequence similarity. Xist Repeats B, C, and D were most similar to each other and to Rsx Repeat 1, whereas Xist Repeats A and E were most similar to each other and to Rsx Repeats 2, 3, and 4. Similarities at the level of k-mers corresponded to domain-specific enrichment of protein-binding motifs. Within individual domains, protein-binding motifs were often enriched to extreme levels. Our data support the hypothesis that Xist and Rsx encode similar functions through different spatial arrangements of functionally analogous protein-binding domains. We propose that the two clusters of repeat domains in Xist and Rsx function in part to cooperatively recruit PRC1 and PRC2 to chromatin. The physical manner in which these domains engage with protein cofactors may be just as critical to the function of the domains as the protein cofactors themselves. The general approaches we outline in this report should prove useful in the study of any set of RNAs.


Assuntos
Marsupiais/genética , RNA Longo não Codificante/química , RNA Longo não Codificante/genética , Animais , Análise por Conglomerados , Humanos , Marsupiais/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Domínios Proteicos , Homologia de Sequência do Ácido Nucleico , Sequências de Repetição em Tandem , Inativação do Cromossomo X
3.
Nat Commun ; 10(1): 2133, 2019 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-31086177

RESUMO

Polycomb (PcG) and Trithorax (TrxG) group proteins give stable epigenetic memory of silent and active gene expression states, but also allow poised states in pluripotent cells. Here we systematically address the relationship between poised, active and silent chromatin, by integrating 73 publications on PcG/TrxG biochemistry into a mathematical model comprising 144 nucleosome modification states and 8 enzymatic reactions. Our model predicts that poised chromatin is bistable and not bivalent. Bivalent chromatin, containing opposing active and silent modifications, is present as an unstable background population in all system states, and different subtypes co-occur with active and silent chromatin. In contrast, bistability, in which the system switches frequently between stable active and silent states, occurs under a wide range of conditions at the transition between monostable active and silent system states. By proposing that bistability and not bivalency is associated with poised chromatin, this work has implications for understanding the molecular nature of pluripotency.


Assuntos
Cromatina/metabolismo , Epigênese Genética/fisiologia , Modelos Biológicos , Proteínas do Grupo Polycomb/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Drosophila/metabolismo , Enzimas/metabolismo , Código das Histonas/fisiologia , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Proteína de Leucina Linfoide-Mieloide/metabolismo , Nucleossomos/metabolismo
4.
Nat Commun ; 10(1): 1874, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015400

RESUMO

Cancer evolution is fueled by epigenetic as well as genetic diversity. In chronic lymphocytic leukemia (CLL), intra-tumoral DNA methylation (DNAme) heterogeneity empowers evolution. Here, to comprehensively study the epigenetic dimension of cancer evolution, we integrate DNAme analysis with histone modification mapping and single cell analyses of RNA expression and DNAme in 22 primary CLL and 13 healthy donor B lymphocyte samples. Our data reveal corrupted coherence across different layers of the CLL epigenome. This manifests in decreased mutual information across epigenetic modifications and gene expression attributed to cell-to-cell heterogeneity. Disrupted epigenetic-transcriptional coordination in CLL is also reflected in the dysregulation of the transcriptional output as a function of the combinatorial chromatin states, including incomplete Polycomb-mediated gene silencing. Notably, we observe unexpected co-mapping of typically mutually exclusive activating and repressing histone modifications, suggestive of intra-tumoral epigenetic diversity. Thus, CLL epigenetic diversification leads to decreased coordination across layers of epigenetic information, likely reflecting an admixture of cells with diverging cellular identities.


Assuntos
Linfócitos B/metabolismo , Cromatina/metabolismo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Leucemia Linfocítica Crônica de Células B/genética , Metilação de DNA , Evolução Molecular , Inativação Gênica , Genes de Cadeia Pesada de Imunoglobulina/genética , Voluntários Saudáveis , Código das Histonas/genética , Histonas/genética , Histonas/metabolismo , Humanos , Leucemia Linfocítica Crônica de Células B/sangue , Proteínas do Grupo Polycomb/genética , Proteínas do Grupo Polycomb/metabolismo , Regiões Promotoras Genéticas/genética , Análise de Sequência de RNA , Análise de Célula Única/métodos , Sequenciamento Completo do Exoma
5.
Nat Commun ; 10(1): 1897, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015486

RESUMO

The cellular decision regarding whether to undergo proliferation or death is made at the restriction (R)-point, which is disrupted in nearly all tumors. The identity of the molecular mechanisms that govern the R-point decision is one of the fundamental issues in cell biology. We found that early after mitogenic stimulation, RUNX3 binds to its target loci, where it opens chromatin structure by sequential recruitment of Trithorax group proteins and cell-cycle regulators to drive cells to the R-point. Soon after, RUNX3 closes these loci by recruiting Polycomb repressor complexes, causing the cell to pass through the R-point toward S phase. If the RAS signal is constitutively activated, RUNX3 inhibits cell cycle progression by maintaining R-point-associated genes in an open structure. Our results identify RUNX3 as a pioneer factor for the R-point and reveal the molecular mechanisms by which appropriate chromatin modifiers are selectively recruited to target loci for appropriate R-point decisions.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , Cromatina/química , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Animais , Butadienos/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cromatina/efeitos dos fármacos , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Subunidade alfa 3 de Fator de Ligação ao Core/antagonistas & inibidores , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 4 Dependente de Ciclina/genética , Quinase 4 Dependente de Ciclina/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Células HEK293 , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Imidazóis/farmacologia , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 1/genética , MAP Quinase Quinase 1/metabolismo , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/metabolismo , Nitrilos/farmacologia , Piperazinas/farmacologia , Proteínas do Grupo Polycomb/genética , Proteínas do Grupo Polycomb/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Proteínas ras/genética , Proteínas ras/metabolismo
6.
Zhonghua Yu Fang Yi Xue Za Zhi ; 53(3): 303-308, 2019 Mar 06.
Artigo em Chinês | MEDLINE | ID: mdl-30841672

RESUMO

Objective: To explore the role of hypothalamus Polycomb Group (PcG) gene (Eed, Ezh) methylation in the relationship between bisphenol A (BPA) exposure during pregnancy and premature puberty in female offspring. Methods: A total of 40 pregnant CD-1 mice were randomly and averagely assigned into four groups: control group (corn oil) and low, middle and high BPA-exposed groups (the poisonous doses were 8 mg/kg, 40 mg/kg, and 200 mg/kg, respectively) by random number table method. Each group was administered by gavage from gestational day (GD) 1 to 18. The vaginal opening of female offspring was observed from postnatal day (PND) 21 to 33. All female offsprings were sacrificed, and hypothalamus was remained on the PND 34. The methylation levels of Eed and Ezh in the hypothalamus were measured. The early puberty of CD-1 mice was evaluated by the rate of vaginal opening in advance, initial time of vaginal opening, the first estrus occurrence and vaginal opening days in advance. The path model was used to explore the role of Eed and Ezh gene methylation in the early puberty of female offspring with maternal BPA exposed including the number of days of vaginal opening in advance as a dependent variable and BPA exposure as an independent variable. Results: The rate of vaginal opening on the 28 day in each maternal BPA-exposure group [low, middle and high BPA-exposed groups were 40.00% (29/72), 47.62% (25/53) and 37.84% (20/53), respectively] was higher than that rate in the control group [14.06%(9/64)]. Similarly, the P(50)(P(25), P(75)) values of initial time of vaginal opening in low, middle and high BPA-exposed group were 28 (26, 30), 28 (26, 29), 28 (26, 30) days, respectively and the P(50)(P(25), P(75)) values of the first estrus occurrence in low, middle and high BPA-exposed group were 31 (27, 32), 30 (27, 31), 31 (28, 33) days, respectively, which were earlier than those in the control group [initial time of vaginal opening was 30(28, 31) days, and the first estrus occurrence was 32(30, 33) days] (all P values<0.05). Compared with the control group (the methylation levels of Eed1, Eed2, Ezh2 were 1.47%, 1.26%, 2.56%, respectively), the methylation levels of Eed1 (1.61%-1.82%), Eed2 (1.36%-1.43%) and Ezh2 (2.87%-3.05%) in female offspring were significantly higher in BPA-exposed groups (all P values<0.05). The results of path model analysis showed that BPA had no direct influence on puberty in advance, but had an indirect effect on puberty in advance (indirect effect path coefficient was 0.045 and 0.142, respectively) by mediating methylation of Eed2, and Ezh2. Conclusion: Early puberty in female offspring induced by maternal exposure to BPA during pregnancy through the increased methylation levels of hypothalamus PcG gene (Eed, Ezh) in female offspring.


Assuntos
Compostos Benzidrílicos/efeitos adversos , Hipotálamo/metabolismo , Fenóis/efeitos adversos , Proteínas do Grupo Polycomb/genética , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Puberdade Precoce/induzido quimicamente , Animais , Feminino , Metilação , Camundongos , Proteínas do Grupo Polycomb/metabolismo , Gravidez , Distribuição Aleatória
7.
Biochim Biophys Acta Gene Regul Mech ; 1862(5): 582-597, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30753903

RESUMO

Co-ordinated interplay between Polycomb group (PcG) and Trithorax group (TrxG) of proteins regulate chromatin state and maintain the transcription "off" and "on" state of a gene in higher eukaryotes. Targeting PcG complex to a specific locus is mediated by DNA sequences known as Polycomb response elements. Interestingly, these PREs are also recognized by TrxG proteins to antagonise PcG mediated gene repression. In this study, we have characterised DNA binding property of rice trithorax group factor ULTRAPETALA1 (OsULT1) which has a SAND domain and B-box motif. Chromatin immunoprecipitation assay indicates cold induced enrichment of OsULT1 occupancy and a decrease in H3K27me3 mark in the promoter region of OsDREB1b gene, during transcription activation. OsULT1 binds to the cis motif "GAGAG", and the sequence specificity is contributed mainly by the SAND domain. GAGAG is one of the cis motifs present in PREs that are recognized by Drosophila GAGA factor and Pipsqueak. Thus, binding of OsULT1 to GAGAG motif, along with a decrease in H3K27me3 suggests that OsULT1 antagonises the repressive effect of PcG complex for transcriptional activation of OsDREB1b. Moreover, OsULT1 interacts with rice SET domain-containing methyltransferase TRX1, suggesting OsULT1 is an integral part of plant Trithorax group complex. Furthermore, the increase in ULT1 levels during environmental cues suggests its involvement in the transcriptional regulation of stress responsive genes. Collectively, these results suggest that the antagonistic functions of PcG and TrxG proteins and the mechanism of recruitment of these complexes to target loci are evolutionarily conserved for gene expression regulation across kingdoms.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Oryza/genética , Proteínas de Plantas/metabolismo , Elementos de Resposta , Sítios de Ligação , Temperatura Baixa , DNA de Plantas/química , DNA de Plantas/metabolismo , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Histona Metiltransferases/metabolismo , Motivos de Nucleotídeos , Oryza/enzimologia , Oryza/metabolismo , Proteínas de Plantas/biossíntese , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas do Grupo Polycomb/metabolismo , Regiões Promotoras Genéticas , Domínios Proteicos , Multimerização Proteica , Estresse Fisiológico/genética , Fatores de Transcrição/genética
8.
PLoS Biol ; 17(2): e3000163, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30807568

RESUMO

During central nervous system (CNS) development, genetic programs establish neural stem cells and drive both stem and daughter cell proliferation. However, the prominent anterior expansion of the CNS implies anterior-posterior (A-P) modulation of these programs. In Drosophila, a set of neural stem cell factors acts along the entire A-P axis to establish neural stem cells. Brain expansion results from enhanced stem and daughter cell proliferation, promoted by a Polycomb Group (PcG)->Homeobox (Hox) homeotic network. But how does PcG->Hox modulate neural-stem-cell-factor activity along the A-P axis? We find that the PcG->Hox network creates an A-P expression gradient of neural stem cell factors, thereby driving a gradient of proliferation. PcG mutants can be rescued by misexpression of the neural stem cell factors or by mutation of one single Hox gene. Hence, brain expansion results from anterior enhancement of core neural-stem-cell-factor expression, mediated by PcG repression of brain Hox expression.


Assuntos
Encéfalo/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster/metabolismo , Histona-Lisina N-Metiltransferase/genética , Proteínas de Homeodomínio/genética , Células-Tronco Neurais/metabolismo , Proteínas do Grupo Polycomb/genética , Fator de Células-Tronco/genética , Animais , Encéfalo/crescimento & desenvolvimento , Proliferação de Células , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Embrião não Mamífero , Regulação da Expressão Gênica no Desenvolvimento , Histona-Lisina N-Metiltransferase/metabolismo , Proteínas de Homeodomínio/metabolismo , Mutação , Células-Tronco Neurais/citologia , Neurogênese/genética , Proteínas do Grupo Polycomb/metabolismo , Transdução de Sinais , Fator de Células-Tronco/metabolismo
9.
Nat Commun ; 10(1): 30, 2019 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-30604745

RESUMO

The inactive X chromosome (Xi) in female mammals adopts an atypical higher-order chromatin structure, manifested as a global loss of local topologically associated domains (TADs), A/B compartments and formation of two mega-domains. Here we demonstrate that the non-canonical SMC family protein, SmcHD1, which is important for gene silencing on Xi, contributes to this unique chromosome architecture. Specifically, allelic mapping of the transcriptome and epigenome in SmcHD1 mutant cells reveals the appearance of sub-megabase domains defined by gene activation, CpG hypermethylation and depletion of Polycomb-mediated H3K27me3. These domains, which correlate with sites of SmcHD1 enrichment on Xi in wild-type cells, additionally adopt features of active X chromosome higher-order chromosome architecture, including A/B compartments and partial restoration of TAD boundaries. Xi chromosome architecture changes also occurred following SmcHD1 knockout in a somatic cell model, but in this case, independent of Xi gene derepression. We conclude that SmcHD1 is a key factor in defining the unique chromosome architecture of Xi.


Assuntos
Proteínas Cromossômicas não Histona/genética , Metilação de DNA/genética , Ativação Transcricional/genética , Inativação do Cromossomo X , Alelos , Animais , Sistemas CRISPR-Cas , Linhagem Celular , Proteínas Cromossômicas não Histona/metabolismo , Ilhas de CpG , Éxons/genética , Feminino , Fibroblastos , Técnicas de Inativação de Genes , Histonas/genética , Histonas/metabolismo , Masculino , Camundongos , Mutação Puntual , Proteínas do Grupo Polycomb/metabolismo
10.
RNA Biol ; 16(2): 176-184, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30608221

RESUMO

Polycomb repressive complex 2 (PRC2) and its methylation of histone 3 at lysine 27 (H3K27me3) play a crucial role in epigenetic regulation of normal development and malignancy. Several factors regulate the recruitment of PRC2 and affects its chromatin modification function. Over the past years, emerging discoveries have portrayed the association of RNA (protein-coding and non-coding) with PRC2 as a critical factor in understanding PRC2 function. With PRC2 being a macromolecular complex of interest in development and diseases, further studies are needed to relate the rapidly evolving PRC2:RNA biology in that scenario. In this review, we summarize the current understanding of different modes of RNA binding by PRC2, and further discuss perspectives, key questions and therapeutic applications of PRC2 binding to RNAs.


Assuntos
Regulação da Expressão Gênica , Proteínas do Grupo Polycomb/metabolismo , RNA/genética , Animais , Humanos , Modelos Biológicos , Proteínas do Grupo Polycomb/genética , Ligação Proteica , RNA/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Pequeno RNA não Traduzido/genética , Pequeno RNA não Traduzido/metabolismo
11.
Cell ; 176(1-2): 182-197.e23, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30595450

RESUMO

During development, the precise relationships between transcription and chromatin modifications often remain unclear. We use the X chromosome inactivation (XCI) paradigm to explore the implication of chromatin changes in gene silencing. Using female mouse embryonic stem cells, we initiate XCI by inducing Xist and then monitor the temporal changes in transcription and chromatin by allele-specific profiling. This reveals histone deacetylation and H2AK119 ubiquitination as the earliest chromatin alterations during XCI. We show that HDAC3 is pre-bound on the X chromosome and that, upon Xist coating, its activity is required for efficient gene silencing. We also reveal that first PRC1-associated H2AK119Ub and then PRC2-associated H3K27me3 accumulate initially at large intergenic domains that can then spread into genes only in the context of histone deacetylation and gene silencing. Our results reveal the hierarchy of chromatin events during the initiation of XCI and identify key roles for chromatin in the early steps of transcriptional silencing.


Assuntos
Cromatina/metabolismo , Inativação do Cromossomo X/genética , Inativação do Cromossomo X/fisiologia , Acetilação , Animais , Cromatina/genética , Células-Tronco Embrionárias , Epigenômica/métodos , Feminino , Inativação Gênica , Histona Desacetilases/metabolismo , Histonas/metabolismo , Camundongos , Proteínas do Grupo Polycomb/metabolismo , Processamento de Proteína Pós-Traducional , RNA Longo não Codificante/metabolismo , Transcrição Genética , Ubiquitinação , Cromossomo X/metabolismo
12.
Cancer Cell ; 35(1): 95-110.e8, 2019 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-30595504

RESUMO

Biallelic inactivation of SMARCB1, encoding a member of the SWI/SNF chromatin remodeling complex, is the hallmark genetic aberration of atypical teratoid rhabdoid tumors (ATRT). Here, we report how loss of SMARCB1 affects the epigenome in these tumors. Using chromatin immunoprecipitation sequencing (ChIP-seq) on primary tumors for a series of active and repressive histone marks, we identified the chromatin states differentially represented in ATRTs compared with other brain tumors and non-neoplastic brain. Re-expression of SMARCB1 in ATRT cell lines enabled confirmation of our genome-wide findings for the chromatin states. Additional generation of ChIP-seq data for SWI/SNF and Polycomb group proteins and the transcriptional repressor protein REST determined differential dependencies of SWI/SNF and Polycomb complexes in regulation of diverse gene sets in ATRTs.


Assuntos
Cromatina/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Proteínas Repressoras/metabolismo , Tumor Rabdoide/metabolismo , Proteína SMARCB1/metabolismo , Teratoma/metabolismo , Sítios de Ligação , Encéfalo/metabolismo , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Epigenômica/métodos , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Humanos , Proteína SMARCB1/química , Análise de Sequência de DNA , Análise de Sobrevida
13.
Nat Commun ; 9(1): 5375, 2018 12 18.
Artigo em Inglês | MEDLINE | ID: mdl-30560907

RESUMO

During development, it is unclear if lineage-fated cells derive from multilineage-primed progenitors and whether active mechanisms operate to restrict cell fate. Here we investigate how mesoderm specifies into blood-fated cells. We document temporally restricted co-expression of blood (Scl/Tal1), cardiac (Mesp1) and paraxial (Tbx6) lineage-affiliated transcription factors in single cells, at the onset of blood specification, supporting the existence of common progenitors. At the same time-restricted stage, absence of SCL results in expansion of cardiac/paraxial cell populations and increased cardiac/paraxial gene expression, suggesting active suppression of alternative fates. Indeed, SCL normally activates expression of co-repressor ETO2 and Polycomb-PRC1 subunits (RYBP, PCGF5) and maintains levels of Polycomb-associated histone marks (H2AK119ub/H3K27me3). Genome-wide analyses reveal ETO2 and RYBP co-occupy most SCL target genes, including cardiac/paraxial loci. Reduction of Eto2 or Rybp expression mimics Scl-null cardiac phenotype. Therefore, SCL-mediated transcriptional repression prevents mis-specification of blood-fated cells, establishing active repression as central to fate determination processes.


Assuntos
Linhagem da Célula/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Proteína 1 de Leucemia Linfocítica Aguda de Células T/metabolismo , Fatores de Transcrição/metabolismo , Animais , Diferenciação Celular/fisiologia , Linhagem Celular , Separação Celular/métodos , Embrião de Mamíferos , Citometria de Fluxo/métodos , Código das Histonas/fisiologia , Mesoderma/citologia , Mesoderma/fisiologia , Camundongos , Células-Tronco Embrionárias Murinas , Proteínas Nucleares/genética , Complexo Repressor Polycomb 1/metabolismo , Proteínas do Grupo Polycomb/metabolismo , RNA Interferente Pequeno/metabolismo , Proteínas Repressoras/genética , Proteína 1 de Leucemia Linfocítica Aguda de Células T/genética , Fatores de Transcrição/genética
14.
Dev Cell ; 47(6): 758-772.e5, 2018 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-30562514

RESUMO

Polycomb repressive complex (PRC) 1 maintains developmental genes in a poised state through monoubiquitination (Ub) of histone H2A. Although Ub-independent functions of PRC1 have also been suggested, it has remained unclear whether Ub-dependent and -independent functions of PRC1 operate differentially in a developmental context. Here, we show that the E3 ubiquitin ligase activity of Ring1B, a core component of PRC1, is necessary for the temporary repression of key neuronal genes in neurogenic (early-stage) neural stem or progenitor cells (NPCs) but is dispensable for the persistent repression of these genes associated with the loss of neurogenic potential in astrogliogenic (late-stage) NPCs. Our results also suggest that histone deacetylase (HDAC) activity of the NuRD/MBD3 complex and Phc2-dependent PRC1 clustering are necessary for the transition from the Ub-dependent to -independent function of PRC1. Together, these results indicate that Ub-independent mode of repression by PRC1 plays a key role in mammalian development during cell fate restriction.


Assuntos
Neocórtex/embriologia , Neocórtex/metabolismo , Complexo Repressor Polycomb 1/metabolismo , Animais , Diferenciação Celular/fisiologia , Linhagem da Célula , Proteínas de Ligação a DNA/metabolismo , Histonas/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Neocórtex/citologia , Proteínas do Grupo Polycomb/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação
15.
Proc Natl Acad Sci U S A ; 115(34): E7987-E7996, 2018 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-30072429

RESUMO

Genome organization plays a fundamental role in the gene-expression programs of numerous cell types, but determinants of higher-order genome organization are poorly understood. In the developing mouse retina, rod photoreceptors represent a good model to study this question. They undergo a process called "chromatin inversion" during differentiation, in which, as opposed to classic nuclear organization, heterochromatin becomes localized to the center of the nucleus and euchromatin is restricted to the periphery. While previous studies showed that the lamin B receptor participates in this process, the molecular mechanisms regulating lamina function during differentiation remain elusive. Here, using conditional genetics, we show that the zinc finger transcription factor Casz1 is required to establish and maintain the inverted chromatin organization of rod photoreceptors and to safeguard their gene-expression profile and long-term survival. At the mechanistic level, we show that Casz1 interacts with the polycomb repressor complex in a splice variant-specific manner and that both are required to suppress the expression of the nuclear envelope intermediate filament lamin A/C in rods. Lamin A is in turn sufficient to regulate heterochromatin organization and nuclear position. Furthermore, we show that Casz1 is sufficient to expand and centralize the heterochromatin of fibroblasts, suggesting a general role for Casz1 in nuclear organization. Together, these data support a model in which Casz1 cooperates with polycomb to control rod genome organization, in part by silencing lamin A/C.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Heterocromatina/metabolismo , Lamina Tipo A/metabolismo , Modelos Biológicos , Proteínas do Grupo Polycomb/metabolismo , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Fatores de Transcrição/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Inativação Gênica/fisiologia , Heterocromatina/genética , Lamina Tipo A/genética , Camundongos , Camundongos Transgênicos , Proteínas do Grupo Polycomb/genética , Células Fotorreceptoras Retinianas Bastonetes/citologia , Fatores de Transcrição/genética
16.
Genes Dev ; 32(11-12): 794-805, 2018 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-29891558

RESUMO

Polycomb-repressive complex 2 (PRC2) is a histone methyltransferase that promotes epigenetic gene silencing, but the dynamics of its interactions with chromatin are largely unknown. Here we quantitatively measured the binding of PRC2 to chromatin in human cancer cells. Genome editing of a HaloTag into the endogenous EZH2 and SUZ12 loci and single-particle tracking revealed that ∼80% of PRC2 rapidly diffuses through the nucleus, while ∼20% is chromatin-bound. Short-term treatment with a small molecule inhibitor of the EED-H3K27me3 interaction had no immediate effect on the chromatin residence time of PRC2. In contrast, separation-of-function mutants of SUZ12, which still form the core PRC2 complex but cannot bind accessory proteins, revealed a major contribution of AEBP2 and PCL homolog proteins to chromatin binding. We therefore quantified the dynamics of this chromatin-modifying complex in living cells and separated the contributions of H3K27me3 histone marks and various PRC2 subunits to recruitment of PRC2 to chromatin.


Assuntos
Cromatina/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Subunidades Proteicas/metabolismo , Linhagem Celular Tumoral , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Edição de Genes , Células HEK293 , Humanos , Indanos/farmacologia , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Proteínas do Grupo Polycomb/antagonistas & inibidores , Ligação Proteica/efeitos dos fármacos , Proteínas Recombinantes de Fusão/metabolismo , Sulfonamidas/farmacologia
17.
Epigenetics Chromatin ; 11(1): 27, 2018 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-29871666

RESUMO

BACKGROUND: Tracking dynamic protein-chromatin interactions in vivo is key to unravel transcriptional and epigenetic transitions in development and disease. However, limited availability and heterogeneous tissue composition of in vivo source material impose challenges on many experimental approaches. RESULTS: Here we adapt cell-type-specific DamID-seq profiling for use in Drosophila imaginal discs and make FLP/FRT-based induction accessible to GAL driver-mediated targeting of specific cell lineages. In a proof-of-principle approach, we utilize ubiquitous DamID expression to describe dynamic transitions of Polycomb-binding sites during wing imaginal disc development and in a scrib tumorigenesis model. We identify Atf3 and Ets21C as novel Polycomb target genes involved in scrib tumorigenesis and suggest that target gene regulation by Atf3 and AP-1 transcription factors, as well as modulation of insulator function, plays crucial roles in dynamic Polycomb-binding at target sites. We establish these findings by DamID-seq analysis of wing imaginal disc samples derived from 10 larvae. CONCLUSIONS: Our study opens avenues for robust profiling of small cell population in imaginal discs in vivo and provides insights into epigenetic changes underlying transcriptional responses to tumorigenic transformation.


Assuntos
Transformação Celular Neoplásica/genética , Proteínas de Drosophila/genética , Drosophila/genética , Discos Imaginais/crescimento & desenvolvimento , Animais , Sítios de Ligação , Cromatina/metabolismo , Imunoprecipitação da Cromatina , Drosophila/embriologia , Drosophila/metabolismo , Proteínas de Drosophila/química , Proteínas de Drosophila/metabolismo , Discos Imaginais/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Ligação Proteica , Análise de Sequência de DNA/métodos
18.
Nat Commun ; 9(1): 1463, 2018 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-29765032

RESUMO

Polycomb repressive complex 1 (PRC1) is an important regulator of gene expression and development. PRC1 contains the E3 ligases RING1A/B, which monoubiquitinate lysine 119 at histone H2A (H2AK119ub1), and has been sub-classified into six major complexes based on the presence of a PCGF subunit. Here, we report that PCGF5, one of six PCGF paralogs, is an important requirement in the differentiation of mouse embryonic stem cells (mESCs) towards a neural cell fate. Although PCGF5 is not required for mESC self-renewal, its loss blocks mESC neural differentiation by activating the SMAD2/TGF-ß signaling pathway. PCGF5 loss-of-function impairs the reduction of H2AK119ub1 and H3K27me3 around neural specific genes and keeps them repressed. Our results suggest that PCGF5 might function as both a repressor for SMAD2/TGF-ß signaling pathway and a facilitator for neural differentiation. Together, our findings reveal a critical context-specific function for PCGF5 in directing PRC1 to control cell fate.


Assuntos
Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Linhagem Celular , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Inativação de Genes , Histonas/metabolismo , Humanos , Camundongos , Neurogênese/genética , Neurogênese/fisiologia , Complexo Repressor Polycomb 1/metabolismo , Proteínas do Grupo Polycomb/deficiência , Proteínas do Grupo Polycomb/genética , Transdução de Sinais , Proteína Smad2/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Ubiquitina-Proteína Ligases/metabolismo
19.
Proc Natl Acad Sci U S A ; 115(19): 4957-4962, 2018 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-29686098

RESUMO

Repressive H3K27me3 and active H3K4me2/3 together form bivalent chromatin domains, molecular hallmarks of developmental potential. In the male germline, these domains are thought to persist into sperm to establish totipotency in the next generation. However, it remains unknown how H3K27me3 is established on specific targets in the male germline. Here, we demonstrate that a germline-specific Polycomb protein, SCML2, binds to H3K4me2/3-rich hypomethylated promoters in undifferentiated spermatogonia to facilitate H3K27me3. Thus, SCML2 establishes bivalent domains in the male germline of mice. SCML2 regulates two major classes of bivalent domains: Class I domains are established on developmental regulator genes that are silent throughout spermatogenesis, while class II domains are established on somatic genes silenced during late spermatogenesis. We propose that SCML2-dependent H3K27me3 in the male germline prepares the expression of developmental regulator and somatic genes in embryonic development.


Assuntos
Histonas/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Regiões Promotoras Genéticas , Espermatogênese/fisiologia , Espermatogônias/metabolismo , Animais , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Histonas/genética , Masculino , Camundongos , Camundongos Knockout , Proteínas do Grupo Polycomb/genética , Espermatogônias/citologia
20.
PLoS Genet ; 14(4): e1007295, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29702639

RESUMO

To better understand the tissue-specific regulation of chromatin state in cell-fate determination and animal development, we defined the tissue-specific expression of all 36 C. elegans presumptive lysine methyltransferase (KMT) genes using single-molecule fluorescence in situ hybridization (smFISH). Most KMTs were expressed in only one or two tissues. The germline was the tissue with the broadest KMT expression. We found that the germline-expressed C. elegans protein SET-17, which has a SET domain similar to that of the PRDM9 and PRDM7 SET-domain proteins, promotes fertility by regulating gene expression in primary spermatocytes. SET-17 drives the transcription of spermatocyte-specific genes from four genomic clusters to promote spermatid development. SET-17 is concentrated in stable chromatin-associated nuclear foci at actively transcribed msp (major sperm protein) gene clusters, which we term msp locus bodies. Our results reveal the function of a PRDM9/7-family SET-domain protein in spermatocyte transcription. We propose that the spatial intranuclear organization of chromatin factors might be a conserved mechanism in tissue-specific control of transcription.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Cromatina/metabolismo , Fertilidade/genética , Histona-Lisina N-Metiltransferase/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Espermatócitos/metabolismo , Animais , Caenorhabditis elegans/enzimologia , Caenorhabditis elegans/fisiologia , Proteínas de Caenorhabditis elegans/genética , Regulação da Expressão Gênica , Células Germinativas , Histona-Lisina N-Metiltransferase/genética , Hibridização in Situ Fluorescente , Masculino , Família Multigênica , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas do Grupo Polycomb/genética , RNA Mensageiro/genética , Transcrição Genética
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